Abstract One of the fundamental questions in biology and by extension, stem cell-based regenerative medicine, is how cell fate is determined in multicellular organisms. Both genetic pathways and epigenetic programs play key roles in lineage specification during the embryonic development of vertebrates. As an important advance, we recently discovered an epigenetic network constituted by a large family of complexes formed by Polycomb group (PcG) proteins. PcG proteins form two major groups of Polycomb repressive complexes (PRCs), PRC1 and PRC2. Even though PRC1 and PRC2 are generally involved in gene repression, mammalian components of PRC1 complexes are exceptionally diverse and their functions are poorly understood. Despite substantial evidence supporting the involvement of PRC1 in cell fate determination, little is known about how the family of mammalian PRC1 complexes coordinates its regulation of lineage specification. This is largely due to the great heterogeneity inherent in the composition of PRC1 complexes in mammals. We propose two independent yet complementary projects to resolve this knowledge gap. Project 1 will elucidate the roles of mammalian PRC1 complexes in cell identity determination. We will utilize a novel approach recently developed by our team to trace the compositional changes of PRC1 complexes in a lineage specific manner and combine this with proteomic, biochemical, and genomic studies. Project 2 will reveal the molecular mechanisms underlying PRC1 complex-mediated transcriptional regulation. This will be accomplished through biochemical analyses and the targeting of individual PRC1 complexes in cells. Taken together, our studies will generate important mechanistic insights into the epigenetic regulation of cell identity, an essential requirement for improving stem- cell based regenerative therapeutic techniques. These approaches also will provide new opportunities to understand the functional diversity of protein complexes in a wide variety of biological processes.